The objective of this proposal is to characterize a novel component of the Neisseria gonorrhoeae oxidative stress and innate immunity defense responses. This proposal is relevant to the mission of the NIH in that N. gonorrhoeae is responsible for a significant health burden in this country and that it addresses a fundamental aspect of N. gonorrhoeae biology that has implications in the pathogenesis of the organism. N. gonorrhoeae is an ideal candidate for studying bacterial oxidative resistance mechanisms due to its obligate association with the oxidative environment of the human host. Preliminary experiments have identified a hydrogen peroxide-induced gene, NG02115, that when mutated confers enhanced resistance of N. gonorrhoeae to hydrogen peroxide, indicating a role for this protein in the oxidative stress response. The NG02115 transcript encodes a putative transcriptional regulator that is hypothesized to control expression of target genes that function in the oxidative stress response. The major focus of Aim 1 will be to determine how NG02115 controls resistance or sensitivity to oxidants by identifying NG02115 target genes. This goal will be accomplished using quantitative RT-PCR and transcriptional microarray techniques. A second goal of Aim 1 is to investigate species-specific differences between the N. gonorrhoeae and Neisseria meningitidis NG02115 coding sequences that may have functional consequences. The overall objective of Aim 2 is to characterize the contribution of NG02115 and its target genes to the N. gonorrhoeae oxidative stress response. Strains harboring mutations in NG02115 and NG02115 regulatory targets will be tested using in vitro killing assays for sensitivity to several types of reactive oxygen species. The amount of macromolecular oxidative damage that accumulates in these strains will also be determined by quantifying oxidative DNA and protein modifications. Together, these experiments will identify the specific types of oxidative stress that NG02115 and its target genes respond to. The final objective of Aim 2 will be to determine the contribution of NG02115 and NG0215 target genes in facilitating either resistance or sensitivity to primary human neutrophils. These neutrophil susceptibility experiments will establish the contribution of NG02115 and its target genes to interactions with a physiologically relevant component of the innate immune system. Relevance: Gonorrhea is a sexually transmitted infection that is a major contributor to the public health burden in the United States. This proposal aims to characterize a novel mechanism by which the causative agent. Neisseria gonorrhoeae survives oxidative stress and killing by innate immune cells and thus is able to promote disease within the human host.